Feline infectious anemia (FIA) is a serious, and sometimes fatal, feline disease with worldwide distribution. The frequency of the disease has been difficult to estimate, due to difficulties detecting the causative agent, Haemobartonella felis. One study estimated that the causative agent was present in 7.5% of the ill cats studied and 3.6% of cats which appeared healthy, while another study reported infection rates as high as 23.2% in animals which had been referred for treatment. (Grindem, C. B. et al., "Risk Factors for Haemobartonella felis Infection in Cats," JAVAM 196:96 (1990); Nash, A. S. and Bobade, P. A. , "Haemobartonella felis Infection in Cats from the Glasgow Area," The Veterinary Record, Oct. 11, 1986, page 373). Cats having low packed cell volume (PCV), a lack of vaccinations generally and against feline enteritis and respiratory tract virus infections specifically, catbite abscesses, a history of roaming outdoors and Feline Leukemia Virus (FeLV) positive status are at a greater risk for FIA. (Grindem et al., supra) Some studies have found a correlation between gender and risk of FIA, although other studies have failed to detect this association. (Hayes, H. M., "Feline Infectious Anaemia. Risk by Age, Sex, and Breed; Prior Disease; Seasonal Occurrence; Mortality," J. Small Anim. Pract. 14: 797 (1973)). Although the disease is curable by treatment with antibiotics, there is no induction of protective immunity. Infection may occur through a variety of routes, including ingestion of infected material, open wounds resulting from fighting, or transplacental transmission.
In addition to severe anemia, clinical manifestations of Haemobartonella felis infection include depression, weakness, anorexia, weight loss, paleness of mucous membranes, and, occasionally, splenomegaly. (Harvey, J. W. "Hemobartonellosis" in Clinical Microbiology and Infectious Diseases of the Dog and Cat, C. E. Greene Ed., W. B. Saunders Co. (1984)). Without intervention, approximately one third of cats with uncomplicated acute haemobartonellosis perish from severe anemia. (Holzworth, J. "Anemia in the Cat," J. Am. Vet. Med. Assoc., 126:471-488, (1956); Splitter, E. J. et al.,"Feline Infectious Anemia," Vet. Med., 51:17-22, (1956)). However, some cats experience both an immune response to the bacteria and a regenerative bone marrow response, which allows erythrocyte production to exceed erythrocyte destruction. These animals eventually recover from the disease, with the recovery time lasting one month or more. (Harvey, J. W. and Gaskin, J. M., "Experimental Feline Haemobartonellosis," J. Am. Anim. Hosp. Assoc., 13:28-38, (1977)).
Unfortunately, cats that recover from acute infections with Haemobartonella felis remain infected with the pathogen for considerable time periods and may even remain infected throughout their lifetimes. (Splitter, E. J. et al. 1956, supra; Harvey, J. W. and Gaskin, J. M., "Feline Haemobartonellosis: Attempts to Induce Relapses of Clinical Disease in Chronically Infected Cats," J. Am. Anim. Hosp. Assoc., 14:453-456, (1978)). In addition, carrier cats may not exhibit any symptoms of infection. The lack of observable symptoms increases the likelihood that the disease will escape treatment and that the carrier will transmit the organism to other animals.
Haemobartonella felis feeds and multiplies on the surface of red blood cells within the circulation, leading to progressive erythrocyte damage and shortened erythrocyte lifespans (Maede, Y., "Studies on Feline Haemobartonellosis. IV. Lifespan of Erythrocytes of Cats Infected with Haemobartonella felis," Jpn. J Vet. Sci., 37:269-272, (1975)). The anemia appears to result from the induction of autoantibodies which result in lysis of the host's own red blood cells. It has been hypothesized that bacterial attachment to the cell surface exposes hidden antigens on the surface or alters antigens normally expressed on the surface. The antibodies produced against the erythrocytes result in a false positive Coombs test.
An alternative hypothesis suggests that cell death is due to antibodies against Haemobartonella felis. It has been proposed that antibodies against Haemobartonella felis cause erythrocyte death by binding to bacteria on the surface of the cell and inducing complement fixation, thereby triggering lysis of the erythrocyte.
Following infection, the levels of bacteria found in the blood may rise and decline rapidly. (Maede, Y. and Hata, R., "Studies on Feline Haemobartonellosis. II. The Mechanism of Anemia Produced by Infection with Haemobartonella felis," Jpn. J Vet. Sci., 37:49-54, (1975); Harvey and Gaskin, (1977), supra). This rapid and extensive fluctuation may be a consequence of splenic sequestration of infected erythrocytes and release of noninfected erythrocytes. (Maede, 1975, supra.).
Prior to the present invention, the most commonly employed method for detecting Haemobartonella felis infection relied on blood film staining. However, this approach suffers from numerous drawbacks. Because the bacteremia associated with Haemobartonella felis infection is episodic, it is often difficult to detect Haemobartonella felis using such traditional cytological approaches. In addition, the pleomorphic nature of Haemobartonella felis may cause the organism to be confused with stain precipitation or cellular debris. (C. B. Grindem et al. supra; C. M. R. Turner et al. Letters The Veterinary Record, Nov. 22, 1986 page 534; H. C. Carney and J. J. England, "Feline Hemobartonellosis," Feline Infectious Diseases 23: 79 (1993)).
Thus, there is a need for improved tests for Haemobartonella felis infection. In particular, given the low levels of Haemobartonella felis present during some stages of the disease, there is a need for sensitive diagnostics capable of detecting the low levels of circulating organisms present during some phases of the infection or in carrier cats harboring low residual levels of the organism. In addition, there is a need for selective tests capable of readily distinguishing Haemobartonella felis from other morphologically or genetically similar organisms. Finally, there is a need for tests which may be rapidly performed. The diagnostics described herein provide all of the preceding advantages.